罗非鱼低氧胁迫转录组响应及耐受性状QTL分析

Global warming causing from climate changes may be one of the biggest challenges to humankind today. Long-term trend of global warming will gradually increase the frequency of occurrence and severity of hypoxia in aquatic systems throughout the world. Recently,the rapid development and application of intensive fish culture system in aquaculture have introduced a strong requirement for fish lines with better hypoxia tolerance trait... The worsening hypoxic conditions could cause serious problems, e.g., direct mortality, reproduction, reduced growth rates and altered behaviour and distributions of aquatic animals, in aquatic ecosystem and aquaculture of commercially important species. The negative effects of hypoxia can be compromised by selection of new fish lines with stronger hypoxia tolerance using molecular marker-assisted selection technique. However,the genetic basis for hypoxia tolerance in fish remains unclear. .. Tilapia is the common name of a group of cichlid fishes native to Africa. Tilapia has become the third most important fish in aquaculture after carp and salmon, with a worldwide production of 3,500,000 metric tons in 2010. To date, no gene expression profiling and quantitative trait locus (QTL) mapping for hypoxia tolerance trait in Tilapia have been reported... Previously we have performed several genomic studies for Tilapia, e.g., in silico mapping of gene-associated SNPs, genome-wide scaning for signatures of recent selection, which provide valuable marker resources for QTL study. This proposed project aims to: (1) identify the genes, pathways and regulation networks involved in response to hypoxia stress in Tilapia by using RNA-seq technique; (2) explore the genetic basis of hypoxia tolerance in Tilapia using a combined strategy,e.g.,candidate gene mapping, QTL linkage mapping and association mapping.The results obtained from this project will deepen our understanding of the adaptations to hypoxia in fish. The availability of QTLs and markers that closely associated to hypoxia tolerance will facilitate the identification of candidate genes in fish by using a map-based cloning method, and will also be a valuable resource for genetic improvement of fish by using marker-assisted selection and/or gene selection methods.

水中溶氧含量不足或缺氧能导致严重的生态和水产养殖问题。应用分子标记辅助育种技术选育具有更强低溶氧耐受能力的鱼类新品种对于提高鱼类低氧逆境胁迫下的生存能力及增加养殖产量具有重要意义。鱼类低溶氧耐受性状属于复杂的经济性状，受到多基因调节。目前鱼类低溶氧耐受性状的遗传基础仍然不清楚。本项目在前期研究基础上，拟首先采用RNA-seq技术对罗非鱼不同器官低溶氧逆境胁迫下的基因时空响应模式进行分析，揭示罗非鱼低溶氧逆境胁迫响应过程中参与调节的功能基因及调控网络；进一步通过结合候选基因作图、数量性状座位（QTL）连锁作图和关联作图等方法，对影响罗非鱼低溶氧耐受性状的遗传网络进行解析，鉴定耐低氧性状相关的QTL及与性状紧密连锁的分子标记。本项目将阐明罗非鱼低溶氧耐受性状的遗传基础，为进一步开展分子标记辅助育种研究奠定基础。

该项目主要是采用转录组学及不同的数量性状作图定位的方法对我国主要养殖的罗非鱼不同品种的低氧逆境胁迫的基因组遗传控制成分进行综合分析，解析低氧耐受性的遗传机理，希望能为罗非鱼耐低氧遗传育种提供基础资料。主要结果如下：.（1）我们首先对罗非鱼鳃组织低氧应激转录组响应进行来分析。共鉴定了239个差异显示基因及34个选择性剪切基因。对选择性剪切基因TLDC2及 SSX2IPA的时空表达分析显示这些基因的剪切与低氧存在显著性的关联。对罗非鱼心脏低氧响应的转录组成分进行了分析，鉴定了大量的差异显示基因。.（2）我们基于本项目产生的数据并从网络数据库中共下载和收集了103个RNAseq数据。这些数据来自于至少15个不同的罗非鱼组织和不同的逆境处理条件，如低氧、高盐、脂肪含量、温度、细菌感染等条件。我们总共鉴定了72,276 高质量的lncRNAs。差异表达分析鉴定了99 个逆境响应相关的lncRNA及1,955组织特异表达的差异表达lncRNA基因。通过以上研究，我们建立罗非鱼不同组织器官低氧逆境响应的基因表达数据库，基本明确逆境响应过程中重要的功能基因及调节网络。.（3）我们首次在罗非鱼基因组上鉴定了4642个拷贝数变异序列（CNV），分析了基因组序列变异与环境（如低氧等）的关系。通过结合GWAS及QTL连锁作图等方法，对罗非鱼群体低溶氧耐受性状遗传控制成份进行解析，共鉴定了4个基因组范围显著的QTL区间，解释的表型效应值为39.5%。功能研究鉴定了两个重要的低氧候选基因GPR132及 ABCG4。群体关联分析在这两个基因外显子上鉴定了多个与低氧耐受性显著相关的SNPs。. 该研究首次在国际上解析了罗非鱼低氧性状的遗传基础，鉴定了重要的低氧性状候选功能基因和分子标记，为进一步开展罗非鱼低氧性状分子育种奠定了良好基础。